The functional gamma-secretase inhibitor prevents production of amyloid beta 1-34 in human and murine cell lines

The amyloid precursor protein (APP) undergoes two consecutive cleavages by different proteases, beta -secretase and gamma -secretase, leading to the r elease of an amyloidogenic 4 kDa fragment called amyloid beta (A beta). Com bining immunoprecipitation and mass spectrometry, we characterized soluble A beta in cultured cell media of mouse neuroblastoma N2a cells and double h APP/hBACE-1 transfected HEK293. The major A beta isoforms detected were A b eta 11-34, A beta1-34, A beta 11-40 and A beta1-40. In this study, we demon strate that overexpression of human beta -secretase (BACE-1) in HEK293 cell s resulted in predominant A beta cleavage at position Glu(11) rather than A sp(1), as well as increased production of A betax-34, but not A betax-40. I ncubation of cells with a specific gamma -secretase inhibitor suggests that cleavage of APP at Leu(34) could be mediated by gamma -secretase itself or by a gamma -secretase dependent process. (C) 2001 Elsevier Science Ltd. Al l rights reserved.